The structure and function of intermediate filaments (IF) are being investigated in a variety of cultured epithelial cells and fibroblasts. In the fibroblast IF system, two major structural proteins (54 kilodalton [desmin, skeletin], and 55 kilodalton [decamin, vimentin]) have been identified and extensively characterized using biochemical and immunological methods. This research program is concentrating on three classes of proteins which coisolate with native fibroblast IF and may represent true IF-associated proteins (IFAPs); one represented by a group of proteins in the 60 to 70 kilodalton molecular weight range and another consisting of a 300 kilodalton molecular weight protein. Three sets of monoclonal antibodies have been prepared which permit us to distinguish the 300 kilodalton, the 60 to 70 kilodalton and the 54/55 kilodalton structural proteins. Preliminary results employing these antibodies in light and electron microscopic studies suggest that the 60 to 70 kilodalton proteins are associated with the nuclear surface and may act to anchor or bind cytoplasmic IF. Recently we have obtained data which suggests that these proteins are very similar to the nuclear lamins. The 300 kilodalton protein appears to be an IF-IF crosslinking protein and recently we have managed to purify it using column chromatographic techniques. In epithelial cells, our efforts are focused on elucidating the function and biochemical properties of IF bundles and their interactions with desmosomes. Studies on desmosome assembly and dynamics are being carried out using cultures of mouse keratinocytes. In addition we are attempting to fractionate desmosomes and to carry out in vitro reconstructions of IF-desmosome complexes. Other studies focus on the relationship between the mitotic spindle and IF. The cell types used in these studies include BHK-21, HeLa, PtK1, mouse epidermal keratinocytes and transformed mouse epidermal cells. A multifaceted approach is taken in all studies and techniques including the isolation and biochemical-biophysical characterization of IF, in vitro polymerization assays, the use of monoclonal and polyclonal antibodies, immunofluorescence and other light optical methods, and all aspects of electron microscopy. The long-range goal of these studies is to determine whether or not IF are directly involved in forming connecting links between the nuclear surface and the plasma membrane. (L)